PROJECT SUMMARY/ABSTRACT At least 50% of patients treated with 125I brachytherapy for uveal melanoma experience significant vision loss within 3-5 years after therapy. Extensive vascular pathology has been reported in the normal retina surrounding the melanoma after 2 years or more. These findings are in line with the accepted, but incompletely tested, concept that microangiopathy causes radiation-related vision loss due to injury of the microvascular endothelium during radiation. This leads to progressive capillary loss after a lag period of several years. If this hypothesis is correct, preventing the development of, or treating, radiation-induced endothelial damage in retinal microvessels will reduce capillary loss and save vision. However, this approach has been difficult to test because early indicators of microvascular endothelial dysfunction have yet to be established. The objectives of the proposed project are to: facilitate earlier detection, treatment and prevention of radiation- associated vision loss. Specifically, we expect to 1. detect early endothelial dysfunction based on reduced blood flow and response to light flicker 2. determine whether it is predictive of subsequent capillary loss and 3. identify molecular mechanisms of radiation-induced endothelial dysfunction. We will apply laser speckle flowgraphy (LSFG) for noninvasive, real-time imaging and measurement of ocular blood flow, in human subjects and mice to test our central hypothesis, that post-radiation endothelial dysfunction is driven by mitochondrial oxidative stress, and is predictive of the severity of subsequent capillary dropout and vision loss. We will test our hypothesis in two aims: Aim 1: Establish whether early impairment of the microvascular endothelial function will predict microvessel drop-out and vision loss in humans after 125I brachytherapy. For this purpose, LSFG, optical coherence tomography (OCT) and OCT-angiography (OCT-A) and tests of visual function will be performed in a prospective cohort of patients undergoing 125I brachytherapy for choroidal melanoma. We will test whether early impairment of ocular blood flow and flicker light-induced vasodilation by LSFG correlate with subsequent vision loss and capillary drop-out (as detected by OCT-A). Aim 2: Test whether selective inhibition of mitoROS production in endothelium prevents the early reduction of blood flow and subsequent loss of capillaries after radiation. In this aim, genetic mouse models in which key regulators of mitochondrial superoxide production are inhibited or overexpressed will be used to test whether inhibition of mitochondrial ROS in the endothelium protects from early impairment of retinal blood flow, endothelial dysfunction and subsequent capillary dropout. The expected outcomes of the proposed studies are knowledge of biomarkers of early radiation retinopathy and insights into the role of endothelial-cell mitochondrial dysfunction in radiation retinopathy. Our studies have the potential to facilitate the development of first-in-class, targeted therapies for radiation-related vision loss.